The effects of progesterone on apoptosis in the human trophoblast-derived HTR-8/SV neo cells
Jin Liu1, Hiroya Matsuo2, Jovelle B. Laoag-Fernandez1, Qin Xu1 and Takeshi Maruo1,3
1Department of Obstetrics and Gynecology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-Cho, Chuo-Ku, Kobe 650-0017, Japan 2Department of Health Sciences, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan
3 Correspondence address. Tel: +81-78-382-6000; Fax: +81-78-382-6019; E-mail: email@example.com
Materials and Methods
Progesterone (P4) is frequently used in the treatment of threatened abortion, prevention of recurrent miscarriage and threatened preterm labor. However, little is known about the molecular mechanism of P4 in the regulation of extravillous trophoblasts’ (EVTs) function. This study was designed to examine the presence of progesterone receptor (PR) in the human trophoblast-derived HTR-8/SV neo cell line, which is a possible model of EVTs, and the effects of P4 on apoptosis in those cells. The HTR-8/SV neo cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum, 100 U/ml penicillin and 100 µg/ml streptomycin. When the cell the population reached 50% confluency, the cells were stepped down to serum-free conditions in the presence or absence of graded concentrations of P4 (1, 10 and 100 ng/ml) for 48 h. The cultured cells were used for RT–PCR, terminal deoxynucleotidyl transferase-mediated 2′-deoxyuridine 5′-triphosphate nick end labeling (TUNEL) assay, immunocytochemistry and western blot analyses. Immunocytochemistry and western blot analyses revealed that PR was evident in HTR-8/SV neo cells. Compared with untreated cultures, treatment with P4 (10 and 100 ng/ml) resulted in significant decreases in the TUNEL-positive rate, Fas, Fas ligand (Fas-L), caspase-8, caspase-3 and poly (ADP-ribose) polymerase (PARP) expression in HTR-8/SV neo cells, and a significant increase in Bcl-2 expression in those cells. Consistently, Fas mRNA expression in those cells was significantly inhibited by the treatment with 10 ng/ml P4 compared with untreated cultures. This study suggests that PR exists in HTR-8/SV neo cells and that P4 inhibits apoptosis by down-regulating Fas, Fas-L, caspase-8, caspase-3 and PARP expression as well as up-regulating Bcl-2 expression in HTR-8/SV neo cells.